Although major depressive disorder (MDD) is heterogeneous and has very low heritability, a genome-wide association study in humans has implicated type III adenylyl cyclase (AC3, adcy3) in MDD. The expression level of human blood AC3 has also been considered as a MDD biomarker, but there is lack of supporting evidence from animal studies. My preliminary data demonstrate that AC3 conventional knockouts exhibit depression-like behaviors and many other related phenotypes including an altered sleeping pattern, supporting AC3 as a candidate depression gene. AC3 is predominantly expressed in primary cilia in the central nervous system (CNS) and is used as a protein marker for primary cilia throughout the brain. However, it is also distributed in other tissues so the pathophysiological role of AC3 in depression is unclear. To overcome the limitation of conventional knockouts and clearly dissect the functions of AC3 in the brain, I have generated an AC3 floxed mouse strain. This mouse strain has been crossed with the EMX1-Cre mouse strain to specifically ablate AC3 in excitatory neurons in the forebrain. It has also been bred wit a tamoxifen-inducible Cre line UBC-Cre/ERT2 in order to knockdown AC3 temporally in adult mice. Moreover, I have prepared a GFP-AC3 lentiviral construct for an AC3-rescue experiment. I will combine molecular biological, morphological, behavioral, electrophysiological and in vivo imaging tools to study the functions of AC3 in the brain. Given that AC3 in olfactory cilia governs the excitation of olfactory sensory neurons, I will first determine if AC3 can similarly regulate neuronal electrical activity in CNS neurons. I will further investigate if AC3 modulates synaptic density and dendritic arborization. Since primary cilia are required for neurogenic Hedgehog signaling, I will test an alternative hypothesis that knockdown of AC3 leads to altered adult hippocampal neurogenesis. In addition, this project will further determine if specifically disrupting AC3 in the forebrain or temporally knockdown of AC3 in adult mice leads to depression-like behaviors in mice. Together, this study could increase our understanding of the functions of AC3 in the brain and has the potential to experimentally consolidate AC3 as a genetic risk factor for major depression.